Initiation of the tumorigenic process can occur when the cellular replication apparatus bypasses an unrepaired DNA lesion resulting in mutation fixation. The objective of the proposed research is to elucidate the basic mechanisms involved in translesion DNA synthesis past radiation damages by studying lesion DNA polymerase interactions both in vitro and in vivo. For this purpose, phage DNA or oligonucleotides containing unique lesions of interest will be prepared as templates for DNA polymerases in vitro. The exact sites of arrest of DNA synthesis or sites of lesion bypass will be determined by analyzing the newly synthesized DNA on sequencing gels. These in vitro results will be correlated with in vivo translesion bypass by measuring the survival of biologically active single stranded DNA containing model DNA damages in uninduced and SOS-induced Escherichia coli. Throughout these studies, the emphasis will be on the structural characteristics of DNA lesions that may affect the stabilization of a nucleotide inserted opposite the lesion through hydrogen bonding and stacking interactions, and on the enzymatic properties of the DNA polymerases used, such as proofreading activity and processivity. Both the properties of the lesions and the properties of the polymerases as they relate to the lesions are important in both mutation fixation and translesion DNA synthesis, since both affect the insertion of a nucleotide opposite the lesion and the efficiency of primer elongation beyond the lesion. Understanding the mechanisms involved in DNA lesion-polymerase interactions should provide a generalized picture of the events that occur when cellular DNA polymerases process potentially carcinogenic radiation- induced DNA damages.